Control of Executioner Caspases with an Allosteric Switch

描述(申请人提供)：蛋白质功能的正交控制允许相关蛋白质家族中的一种蛋白质失活，以便评估其独特的生物学功能。我们建议在caspase家族中开发一种新的正交控制，称为变构开关。半胱氨酸蛋白酶是一种半胱氨酸蛋白酶，它通过切割一系列不连续的目标蛋白来执行细胞死亡，最终导致细胞凋亡。半胱氨酸天冬氨酸氨基转移酶有一个大的中央空腔，可以被小分子变构抑制。Caspase被认为是治疗癌症(caspase激活剂)或中风、心脏病发作和阿尔茨海默病(caspase抑制剂)的良好靶点。不幸的是，目前可用的结合在caspase活性位点上的小分子抑制剂的相似性并不能明确地识别这12种不同的caspase的确切作用。我们将使用定向进化和计算蛋白质设计的组合来在caspase-3、-6和-7中引入变构开关。当我们选择或设计的小分子效应器与进化的变构位点结合时，caspase的活性将被抑制。这一开关将允许我们一次关闭一种类型的caspase，以确定哪些蛋白被caspase-3、-6或-7特异性切割。被裂解的蛋白质本身就很有趣，因为它们的裂解是促凋亡的，这意味着它们会导致细胞死亡。诱导细胞凋亡的能力是一种有用的抗癌药物的标志。我们的研究可能会建议哪种半胱氨酸氨基转移酶应该作为治疗哪种疾病的靶点。这些caspase中变构开关的成功实施将为其他9个caspase中的变构开关铺平道路，并最终进入缺乏生物信息的其他蛋白质家族。 非技术综述：疾病治疗的中心问题是知道哪种蛋白质应该作为治疗哪种疾病的靶点。我们将在半胱氨酸酶中开发的变构开关技术只允许一种类型的蛋白质被药物关闭，以确定通过靶向该特定蛋白质可以有效治疗哪种疾病。我们专注于caspase蛋白，它可以导致癌细胞死亡。

DESCRIPTION (provided by applicant): Orthogonal control of protein function allows one protein from a family of related proteins to be inactivated so that its unique biological function can be assessed. We propose development of a new kind of orthogonal control, called allosteric switches, in the caspase family. Caspases are cysteine proteases that execute cell death by cleaving a discrete selection of target proteins, which ultimately leads to apoptosis. Caspases have a large central cavity that can be allosterically inhibited by small molecules. Caspases are believed to be good targets for treatment of cancer (caspase activators) or stroke, heart attack and Alzheimer's Disease (caspase inhibitors). Unfortunately the similarities in currently available small inhibitors that bind at the caspase active sites have not allowed unambiguous identification of the precise roles of the twelve different caspases. We will use a combination of directed evolution and computational protein design to introduce an allosteric switch in caspases-3, -6, and -7. When the small molecule effector we have selected or designed against binds to the evolved allosteric site, caspase activity will be inhibited. This switch will allow us to turn off one type of caspase at a time to determine what proteins are specifically cleaved by caspase-3 or -6 or -7. The cleaved proteins themselves are inherently interesting, because their cleavage is proapoptotic, meaning they lead to cell death. The ability to induce apoptosis is the hallmark of a useful cancer drug. Our studies may suggest which caspase should be targeted for treating which disease. Successful implementation of allosteric switches in these caspases will pave the way for allosteric switches in the other nine caspases and ultimately into other families of proteins where biological information is lacking. NON-TECHNICAL SUMMARY: The central problem in disease treatment is knowing which protein should be targeted to treat which disease. The allosteric switch technology we will develop in caspases allows just one type of protein to be turned off by a drug, to determine which disease could be effectively treated by targeting that particular protein. We focus on caspase proteins, which can cause cancer cells to die.